MUD transcript: 24-03-00
Course room(s): PPS
23:02:01 ClareS connects.
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23:02:09 ClareS says "Hi everyone"
23:02:36 ClareS says "sorry I am late..."
23:02:45 SarahL says "Hi Clare, how are you tonight"
23:02:59 RobertO says "hello clare hope you are well"
23:03:01 ClareS says "first of all I would like to apologise for the mistake in my last message to the list"
23:03:29 ClareS says "I am organising a conference (in cyberspace) that starts on June 26, that was probably why it was on my mind"
23:03:50 LesleyM connects.
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23:03:55 ClareS says "of course, it is ** this Sunday** that is the start of British Summer Time: March 26"
23:04:01 ChristopherA says "Hi Clare"
23:04:07 ClareS says "it is really good to see so many of you here"
23:04:23 VijayK connects.
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23:04:28 LesleyM says "Hello everyone"
23:06:15 VijayK says "hello again. I am back briefly, was here yesterday"
23:06:16 ClareS says "the main subject of this session will be Molecular Forces -- how are you all getting on with this section?"
23:06:33 LesleyM says "I would like to know how the entropy calculations are done in the Protein solv. interactions"
23:07:05 StephanieD says (to ClareS) "Thanks anyway for the time..ly reminder!"
23:07:19 ClareS is searching for the correct page of course materila in a slow browser
23:07:30 LesleyM says "what measuements are they based on?"
23:07:43 ClareS says (to StephanieD) "yes -- remember that we will announce times of meetings in GMT, whatever the time of year"
23:09:08 ClareS says (to LesleyM) "do you have a reference to a page of the course material?"
23:10:03 ChristopherA says (to MichealS) "Ye I'm not too bad if only my computer would stop playing up"
23:10:51 ClareS says "it is very difficult to calculate entropy"
23:11:12 LesleyM says "ppscore/section7/solvent.html - no numbers are given"
23:11:31 ClareS says "you can think of entropy qualitatively as being equivalent to disorder"
23:11:35 SarahL says "http://pps9900.cryst.bbk.ac.uk/ppscore/section7/solvent.html"
23:12:09 ClareS says "e.g. if you have to remove a (disordered) water molecule from an active site cleft..."
23:12:24 LesleyM says "yes, but than how is it accounted for?"
23:12:35 ClareS says "in order to replace it by a more static drug molecule, that will decrease the overall entropy of the system"
23:12:46 ClareS says "which is generally unfavourable"
23:13:26 ClareS says "in most (if not all.. I'm slightly out of date) modelling programs, entropy is not taken into account"
23:13:33 LesleyM says "how do you decide it is disordered...after all is in cleft site a degree of order is implied."
23:14:29 ClareS says "if you do a calculation called a free energy perturbation the entropy cancels out of the equation and can be ignored"
23:14:46 VijayK says "Is entropy generaly disordered>'"
23:14:59 ClareS says "you do this by comparing two very similar systems with the same (unknown) degree of entropy"
23:15:22 ClareS says (to VijayK) "entropy is a measure of disorder.. remember the 2nd law of thermodynamics"
23:15:37 LesleyM says "waht is the free energy perturbation equation?"
23:16:51 ClareS says (to LesleyM) "it's not an equation so much as a calculation method"
23:16:57 RobertO says "to clares a disordered water then would be one NOT hydrogen bonded to a residue in the cleft?"
23:17:57 ClareS says "you calculate the enthalphy change in moving from one system to another with the same amount of entropy, the entropies cancel out so the difference is the same as the difference in overall energies"
23:18:22 LesleyM says "also - are the water molecules in a crystal tken to be ordered solvent molecules inthe first solvation shell, or are they intrisic to the structure?"
23:18:23 ClareS says (to LesleyM) "is that clear.. I'm afraid I can't remember the detailed maths"
23:19:14 LesleyM says "certainly clearer - it seems to me that quantifiable delta entropy must be important somewhere inthis though"
23:20:05 ClareS says (to RobertO) "yes and no... waters are not either "disordered" or "ordered" but have degrees of disorder"
23:20:43 ClareS says "any water molecule, whether h-bonded to a protein residue or not, will have more freedom of movement than a drug molecule that is tightly bound into the active site"
23:21:20 MichaelS says " regarding Robert's water question, is the degree of disorder related ot things like temperature?"
23:21:30 ClareS says (to LesleyM) "it's surprising that molecular mechanics modelling works so well since it makes very many approximations"
23:22:12 ClareS says "and now back to Lesley's question on water molecules"
23:22:48 ClareS says "almost all water molecules will just be those in the first solvation shell (they are often observable in crystal structures)"
23:23:09 LesleyM says "to MichaeIS delta entropy is equal to delta heat/absolute temp. Yes temp is important."
23:23:14 ClareS says "occasionally a water molecule will be really important for the way (e.g.) an enzyme works"
23:25:07 ClareS says "one example is the aspartic proteases - they have two Asp residues very close together in the active site"
23:25:21 VijayK says "what is meant by delta entropy?'"
23:25:30 ClareS says "one is protonated and they are held in place by a tight h-bonding network including a water molecule"
23:25:36 LesleyM says "are the waters of the solvation shell essentially like those in a smaller hydrate or are they,as it were, looser?"
23:26:09 LesleyM says "delta entropy is change in entropy"
23:26:20 ClareS says (to LesleyM) "thanks for answering Michael's question, my Netscape crashed"
23:27:19 ClareS says (to VijayK) "delta just means a change - delta entropy is a change in entropy"
23:27:43 VijayK says "Thanks "
23:28:05 ClareS says "you might remember using equations with delta x / delta y when learning basic calculus at high school"
23:28:18 VijayK says "Yes I do"
23:28:31 StephanieD says "How can one infer that a water molecule is contributing to function or structure, rather than simply being a solvent molecule?"
23:29:08 ClareS says (to LesleyM) "the waters in a solvation shell are certainly "looser" than those in a hydrate -- an individual molecule will usually move between hydrogen bonding positions over time"
23:29:18 ClareS says (to StephanieD) "it's difficult..."
23:29:51 ClareS says (to StephanieD) "if you have a lot of crystal structures of the same type of protein, you might notice one water in the same position in each"
23:30:19 ClareS says "that would be called an invariant water molecule -- a good sign that it was important for something..."
23:31:20 SarahL says "so in this case the water is contributing to the structure of the protein as it is present all the time eveen in a crystal structure"
23:31:26 MichaelS says (to ClareS) "is that why you use deuterated water in NMR to "see" the side-chain H's?"
23:31:49 StephanieD says "True."
23:32:04 ClareS says "also you might know the mechanism of an enzyme interaction and be able to work out that a water molecule was important"
23:32:37 ClareS says (to MichaelS) "yes -- you can only see water oxygens in X-ray structures"
23:33:00 ClareS says "so you can't know the exact hydrogen bonding pattern"
23:33:37 SarahL says "can you give us an example of where this is true"
23:33:53 RobertO says "unless you have better than 1 angstrom resolution then the water hydrogens and hydrogen bonds are visible"
23:34:01 ClareS says (to SarahL) "there will always be water around a crystal structure, but the waters will often be difficult to see as they are so disordered"
23:34:22 ClareS says (to RobertO) "yes... but there are few of these structures in the database even now"
23:34:44 VijayK says "Iam afraid I ahve to say bye bye now'"
23:34:45 ClareS says (to SarahL) "an example of where *what* is true??"
23:34:56 RobertO says "I have a structure at 1.6A and it shows the hydrogen bonds well for the functionally important waters"
23:35:10 ClareS says (to SarahL) "sorry, it's difficult to keep up with several conversations at once"
23:35:15 SarahL says "the water molecule being part of the structure or function of the protein,i mean sorry"
23:35:35 ClareS says (to RobertO) "that is a very high resolution (accurate) structure -- what is the protein?"
23:35:51 RobertO says "yes very few and not easy to get by any means...sometimes luck"
23:36:00 ClareS says (to SarahL) "I have already mentioned aspartic proteases as a good example of this"
23:36:33 RobertO says "it is a dimeric thymadylate synthase from E. coli"
23:37:17 VijayK disconnects.
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23:37:25 SarahL says "Thank you I have noted that down"
23:37:54 ClareS is trying to think of others from memory
23:38:05 ClareS says (to RobertO) "how many residues?"
23:38:18 StephanieD says "I'm having some difficulty understanding the nature of the hydrophobic bond. What criteria are used to define degree of hydrophobicity? The hydrophobicity scales appear to vary somewhat between themselves."
23:38:58 RobertO looking ambarassed...
23:39:14 ClareS says "that is because the hydrophobic scales are all calculated using slightly different criteria"
23:39:28 RobertO says "I think 470 but cant remember"
23:40:39 ClareS says (to RobertO) "that is quite a large structure to have been solved at such high resolution"
23:41:33 ClareS says "some calculate hydrophobicity from the chemical properties of the sidechains, others measure partition coefficients, etvc"
23:41:59 RobertO says "yes I currentlu hold the record for the highest resolution structure of TS, it was quite unexpected and it is a synchrotron data set"
23:42:17 ClareS is impressed
23:42:19 LarryT says "I got the impression that post Kyte-Doolittle, vatious scales were "adjusted" to correlate new hydrophobicity scales with experimental data for non-globular proteins"
23:42:31 ClareS says (to RobertO) "is that structure in the PDB yet?"
23:43:05 RobertO says "I have the structure of the apo enzyme and am working on the binary and ternary structure with ligands bound...all at 1.6 to 1.8A"
23:43:14 ClareS says (to LarryT) "yes, but no-one can quite agree on the "best" way (if indeed there is one) to adjust them"
23:43:39 ChristopherA says (to ClareS) "Which of the hydrophobicity methods is best or as so often the case, do each have advantages?"
23:43:50 RobertO says "no it will go in in about a month when the paper is done"
23:44:23 ClareS says (to ChristopherA) "there is no one method which is "best" in all circumstances"
23:44:54 ClareS says "(that is, for all types of protein and all types of calculation that use these values)"
23:44:59 SarahL says "that sounds very impressive Robert"
23:45:37 LarryT says "i ASSUME "best" is the one that more closely fits YOUR experimental data (g)"
23:46:39 ClareS says "there are hydrophobicity scales that have been developed specifically for predicting transmembrane helices"
23:47:05 RobertO says (to ClareS) "I get really luck sometimes, TS has been solved and worked on for over 20 years and I just tried some new conditions and it happened"
23:48:03 SarahL disconnects.
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23:48:09 RobertO says (to ClareS) "the real purpose was to see a new drug lead in the active site and look at the interactions to compare real life with a molecular docking run"
23:48:14 ClareS is still impressed ;)
23:49:02 RobertO says "Thanks"
23:49:04 LesleyM says "what is the underlying difference in the scales - dielectric constant...?"
23:50:39 ClareS says (to LesleyM) "you mean hydrophobicity scales?"
23:50:52 MichaelS says "With situations such as Robert describes, and thinking about one PhD project at my first job in Sydney, are crystals made with salts, to mimic reality?"
23:50:59 LesleyM says "yes"
23:51:29 ClareS says (to RobertO) "could you send us at BBK a reprint of the paper when it's out, please?"
23:51:59 ClareS says (to MichaelS) "it is difficult to crystallise proteins without salts"
23:52:39 RobertO says (to ClareS) "absolutely, and I could send coordinates earlier when they are cleaned up and ready for the paper"
23:52:42 ClareS says "that is why people (like Robert ;) can sometimes spend years getting the conditions "just right" before they get good crystals"
23:52:52 SarahL connects.
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23:52:53 ClareS says (to RobertO) "thanks!"
23:53:31 SarahL says "sorry, I got disconected "
23:53:34 MichaelS says "Are ther any particular salts used?"
23:53:55 ClareS says (to LesleyM) "different scales are calculated using different methods -- some are experimentally derived, others theoretical"
23:54:32 ClareS says (to MichaelS) "many different ones: there is a database of protein crystallisation conditions somewhere on the web but I can't remember where"
23:54:51 ChristopherA says (to MichealS) "Yes ammonium sulphate is often used"
23:55:06 MichaelS says "thanks. I'll try a search for it."
23:55:12 ClareS says "of course, some salts are much more often used than others"
23:55:44 ClareS says "try the British Crystallographic Association to start with -- its pages are hosted by our department"
23:56:30 RobertO says "If you are interested you should pick up a Hampton catalog that gives the most common conditions for crystallization of proteins and has kits for screening conditions"
23:57:09 ClareS says (to RobertO) "yes, but the one on the web is free and more easily accessible"
23:57:24 LesleyM says (to RobertO) "does Hampton have a website"
23:57:30 ClareS says "from wherever you are ;)"
23:57:39 RobertO says "Many of the conditions there will yield a crystal of some sort for your protein then you can "tune" the conditions to get a better crystal"
23:57:55 MichaelS says "Thanks Robert, but I think I'll try for the web one, as clare says;-)"
23:58:10 RobertO says "yes hampton does have a web site but I dont know the address"
23:58:14 ClareS says "it is relatively easy to get crystals but often you get crystals that don't diffract or do so very poorly"
23:58:48 MichaelS says "The next place I'm going to, to do my doctorate, is more into NMR, CD etc than crystals, but I might get to do some."
23:58:59 LesleyM says (to RobertO) "thanks"
23:59:21 ClareS says (to MichaelS) "where will you be going? (just out of idle curiosity ;)"
00:00:08 ClareS would like to end the meeting in say 5-10 minutes: are there any burning questions that you'd like to raise before I do so?
00:00:12 MichaelS says (to ClareS) "To Sydney University, start of the year, with Joel Mackay. He's into transcription factors."
00:00:38 RobertO says "the website is www.hamptonresearch.com"
00:00:53 LesleyM says (to ClareS) "no - thank you for all your help."
00:00:58 ClareS says (to MichaelS) ".. 2001? then it shouldn't affect your participation in this course"
00:01:35 LesleyM says (to RobertO) "many thanks Robert"
00:01:46 MichaelS says "Not at all. In fact, I'm using this time to get back into the swing of things."
00:02:44 ClareS looks round expectantly..
00:03:08 LesleyM says "goodbye everyone - interesting conversation!"
00:03:13 SarahL says "when is the next assignment?"
00:03:48 RobertO says "Thanks clare!"
00:03:48 ClareS says "thanks to you all for turning up and for making it such a lively discussion"
00:04:28 StephanieD says "Thanks for the clarifications..."
00:04:50 MichaelS says "Goodnight all."
00:04:51 ClareS says "there will be an assignment and a quiz after the next section (i.e. between sections 8 & 9)"
00:05:15 LesleyM disconnects.
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00:05:42 SarahL says "thankyou, clare it must be time for bed goodnight"
00:05:47 StephanieD says "See you all next time."